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Volume 153, Issue 8

Analysis of the AAA+ chaperone gene and stress-response expression in the halophilic methanogenic archaeon Free

Abstract

ClpB is a member of the protein-disaggregating chaperone machinery belonging to the AAA+ superfamily. This paper describes a new gene from the halophilic methanoarchaeon , which has not been reported previously in Archaea. The partial sequence of was identified from the investigation of the salt-stress response of by differential-display RT-PCR (DDRT-PCR). Furthermore, the complete sequence (2610 nt) and its upstream genes encoding the type I chaperonin GroEL/ES were obtained through inverse PCR, Southern hybridization and sequencing. The G+C ratio of is 49.6 mol%. The predicted ClpB polypeptide contains 869 aa and possesses a long central domain and a predicted distinctly discontinuous coiled-coil motif separating two nucleotide-binding domains (NBD1 and NBD2). NBD1 has a single Walker A and two Walker B motifs and NBD2 has only one of each Walker motif, a characteristic of HSP100 proteins. Two repeated Clp amino-terminal domain motifs (ClpN) were identified in ClpB. The putative amino acid sequence shared 75.6 % identity with the predicted homologue annotated as ATPase AAA-2 of DSM 6242. Preliminary phylogenetic analysis clustered ClpB (ClpB) with the low G+C Gram-positive bacteria. Stress response analysis of by Northern blotting showed up to 1.5-fold increased transcription levels in response to both salt up-shock (from 2.1 to 3.1 M NaCl) and down-shock (from 2.1 to 0.9 M NaCl). Both and transcript levels increased when the temperature was shifted from 37 °C to 55 °C. Under heat stress transcription was repressed by the addition of the osmolyte betaine (1 mM). In conclusion, a novel AAA+ chaperone gene from a halophilic methanogen that responded to the fluctuations in temperature, salt concentration and betaine has been identified and analysed for the first time.

  • Received:
  • Accepted:
  • Revised:
  • Published Online:
Keyword(s): DDRT-PCR, differential-display RT-PCR , HSP, heat-shock protein and NBD, nucleotide-binding domain
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2007-08-01
2024-04-19
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Analysis of the AAA+ chaperone clpB gene and stress-response expression in the halophilic methanogenic archaeon Methanohalophilus portucalensis
Microbiology 153, 2572 (2007); https://doi.org/10.1099/mic.0.2007/007633-0
/content/journal/micro/10.1099/mic.0.2007/007633-0
/content/journal/micro/10.1099/mic.0.2007/007633-0
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